Selected article for: "lysed cell and lysis buffer"
Author: Teoh, Kim-Tat; Siu, Yu-Lam; Chan, Wing-Lim; Schlüter, Marc A.; Liu, Chia-Jen; Peiris, J. S. Malik; Bruzzone, Roberto; Margolis, Benjamin; Nal, Béatrice
Title: The SARS Coronavirus E Protein Interacts with PALS1 and Alters Tight Junction Formation and Epithelial Morphogenesis
  • Document date: 2010_11_15
    • ID: ufw13pjx_20
    Snippet: Forty-eight hours post-transfection with either pcDNA-E (wt), pcDNA-HA-E (wt), pcDNA-HA-E (ΔPBM), or pSectag2B-Myc-CRB3 constructs, cells were lysed with 0.5–1 ml of cell lysis buffer (20 mM Tris-HCl pH7.5, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 1 mM PMSF, 1X Complete Protease Cocktail inhibitor), and centrifuged at 4000 × g for 20 min at 4°C to remove large cellular debris. For GST-pull down assay, 400 μl of cell lysates were incubated o.....
    Document: Forty-eight hours post-transfection with either pcDNA-E (wt), pcDNA-HA-E (wt), pcDNA-HA-E (ΔPBM), or pSectag2B-Myc-CRB3 constructs, cells were lysed with 0.5–1 ml of cell lysis buffer (20 mM Tris-HCl pH7.5, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 1 mM PMSF, 1X Complete Protease Cocktail inhibitor), and centrifuged at 4000 × g for 20 min at 4°C to remove large cellular debris. For GST-pull down assay, 400 μl of cell lysates were incubated overnight at 4°C with purified GST fusion proteins of PALS1 (0.5 μg and 1 μg). Glutathione beads were washed five times with cell lysis buffer by sequential rounds of centrifugation at 4°C. The final pellet containing interacting proteins was analyzed by electrophoresis and immunoblotting.

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